Cytotoxicity Evaluation of Eucalyptus camaldulensis leaf extract against Cholangiocarcinoma Cell lines (CL-6)

Abstract Background Cholangiocarcinoma(CCA) is an important public health problem in many tropical and subtropical parts of the world. Eucalyptus camaldulensis leaves as well as its essential oil extracts are well known for its traditional use and pharmacological actions including insect repellant/insecticidal activity, antimicrobial, cytotoxic/anticancer activity as well as for many other disorders in different parts of the world including Africa and Southeast Asia. Materials and Methods This study was conducted to investigate the cytotoxic activity of E. camaldulensis leaf extract preparations at in vitro against the CCA tumor (CL-6 cell line). E. camaldulensis leaves collected from Ethiopia (Western Gojjam and Southern Gonder area) was used for extract preparation in 95% ethanol using different extraction methods, i.e.,ultrasonication, heating reflux, and maceration (1 day, 3 days and 7 days). The extract and the standard anticancer drug preparation 5-fluorouracil (5-FU) were evaluated in triplicate for their cytotoxic activities against CL-6 cell line and normal human cell line (OUMS) using standard MTT cell proliferation assay at eight serial dilution concentrations (3.8–500µg/mL). Results A semi-solid brown black residue with a characteristic odor was obtained after solvent removal and extract concentration was done by using a rotary evaporator at 60oC. The highest (mean ± SD) percent yield (3.73 ± 0.61) was obtained by using maceration technique with the solvent for 7 days, followed by heating reflux (1.73 ± 0.80) and the least was obtained from 1day maceration method (0.90 ± 0.52). The extract was then stored at -20oC until use. The cytotoxic activity (mean ± SD IC50 values) E. camaldulensis extract and 5-FU against CL-6 were 102.29 ± 5.87 and 106.75 ± 3.06 (mean IC50 ± SD) µg/mL, respectively. The extract’s IC50 value observed in normal OUMS cell line was 165.9 ± 10.3. Conclusion The extract of E. camaldulensis collected from the study area exhibited in vitro cytotoxic activity against human CCA tumor cell line (CL-6). Further in vitro and in vivo evaluation of the selected herbal extract preparation is recommended in the search for development of safe and promising drug options against CCA and other cancer types.